With the aim to measure the relative curvature radius between the two segments in spherical segmented mirrors and to improve the matching accuracy of segments, a new method utilizing a Shack-Hartmann sensor and a high precision spherometer to measure the relative radius of each spherical segment is presented and an experiment system is also setup. The method firstly uses a co-focus technique to co-focus each segment; then uses the S-H sensor to measure the defocus in the optical axis direction and employs a PZT actuator behind the segments to make the defocus near zero. After the same co-focus process is undertaken again, the relative radius of each segment can be measured by the high precision spherometer. In the experiment, the segmented mirror consists of three spherical hexagonal segments with an opposite side of 300 mm and a curvature radius of 2 000 mm. The analysis and experiment indicate that the precision of the method is about 1 μm, which shows that the method is suitable for the measurement of relative curvature radius of large spherical segmented mirrors.